Natural antibodies, either polyclonal or monoclonal, have been used as specific binding reagents for a considerable time. When immobilised on solid phases they can be used in purification procedures.
Antibodies are large complex multi-chain proteinaceous structures. Although it has been appreciated for some while that substantial portions of these structures seem unrelated to the specific binding properties of the antibodies, the minimum portion necessary to provide adequate specific binding has been a matter of debate. It has already been shown that so-called Fv fragments, ie. an antibody fragment essentially comprising only a single heavy-chain variable region and its corresponding light chain variable region, can exhibit specific binding activity. Very recently it has also been shown by Ward et al (Nature, 1989, Vol. 341, p544-546) that a single variable domain from an antibody ("Dab") can exhibit significant specific binding activity. The production of single variable domain antibodies (Dabs), as described by Ward et al, is also described in detail in EP 0368684 A1 (Medical Research Council) published on May 16, 1990. There is now some evidence that peptides much shorter than the Dab, so-called paralogs, can be designed to mimic antibody binding to some extent (Kauva et al, Biochromatography, 5, 1990, p22).
To be of practical use in immunoadsorption processes, specific binding activity alone is not sufficient. The specific binding agent must also be capable of being linked to a solid phase such as a carrier material in a column. Ideally, this linkage is achievable without any significant adverse affect on the specific binding activity. Such adverse affects can easily arise through chemical or conformational changes in the specific binding region, or simply by physical (steric) hindrance of access to the specific binding region. In the case of conventional specific binding reagents, particularly whole antibody molecules or large portions of such molecules such as Fab fragments, the specific binding region or regions comprise only a minor proportion of the total molecule. The comparatively vast residual bulk of the molecule, which is apparently not directly involved in the specific binding activity, provides abundant scope for the existence of locations which can participate in chemical or physical linkage to solid phases. These regions can be relatively remote from the essential specific binding regions, such that the resulting linkages need not interfere with the specific binding activity.
However, in the case of a specific binding entity essentially comprising only one or more variable domains unassociated with any substantial portion of the originating antibody or antibodies, eg. a Fv fragment or a single variable domain, the relative proportion of the molecule which participates in the essential specific binding activity is very much higher. Indeed, it might be expected that any attempt to link the small specific binding entity to a solid phase would entail a very high risk that the essential specific binding activity will be adversely affected.